Atherosclerosis is a progressive inflammatory disease and the underlying cause of heart attack and stroke. Macrophages play a crucial role in the formation and progression of atherosclerotic lesions. Macrophage apoptosis occurs throughout all stages of atherosclerosis with a differential impact on lesion morphology in early versus late atherosclerosis. Loss of macrophages in early lesions is thought to reduce lesion size, whereas cell death in advanced lesions contributes to the necrotic core and plaque destabilization. Studies by Tabas and coworkers have demonstrated that defective phagocytic clearance results in apoptotic cell accumulation in atherosclerotic plaques. Here we propose that the intrinsic ability of macrophages to resist pro- apoptotic stimuli may be another important determinant of macrophage survival and apoptotic cell numbers in atherosclerotic lesions. There are two major pro-survival pathways, PI3K/Akt and NF-kB, and both are constitutively active in macrophages and macrophage-derived foam cells of atherosclerotic lesions. Recent studies in our laboratory have shown that genetic deficiency of the prostaglandin E2 receptor, EP4, in hematopoietic cells promotes macrophage apoptosis in atherosclerotic lesions by modulating the PI3K/Akt and NF-kB signaling pathways. Two Akt isoforms are expressed in macrophages, Akt1 and Akt2, yet their relative contributions to macrophage apoptosis and atherogenesis have not been determined. Interestingly, Akt has been reported to mediate signaling through IKKa that may activate the NF-kB pathways with its anti-apoptotic activity. We hypothesize that cross-talk between the Akt and NF-kB signaling pathways is a critical determinant of macrophage survival and atherogenesis. In this proposal we intend to define the contribution of distinct members of the Akt NF-kB signaling pathways, including Akt1, Akt2, and IKKa, to macrophage survival and atherosclerotic lesion formation. We hypothesize that both Akt-1 and Akt-2 contribute to macrophage survival but that deficiency of both isoforms will promote macrophage apoptosis to a greater extent than deficiency of either isoform alone. The goal of Specific Aim 1 is to examine the impact of hematopoietic cell deficiency of Akt1, Akt2, or both on macrophage survival and atherogenesis in LDLR-/- mice in vivo. The goal of Specific Aim 2 is to define the impact of macrophage deficiency of Akt1 and/or Akt2 on apoptosis and the Akt and NF- :B signaling pathways in vitro. Akt and IKKa are necessary for TORC1 formation in signal transduction. Therefore, we will examine the hypothesis that macrophage deficiency of Akt1 and/or Akt2 will suppress mTOR activity. In Specific Aim 3, we will examine the hypothesis that IKKa deficiency in hematopoietic cells will reduce macrophage survival and impact atherogenesis through alterations in the Akt and NF-:B signaling pathways. A better understanding of the molecular mechanisms of macrophage survival may provide new targets for the prevention of atherosclerosis and cardiovascular events. PUBLIC HEALTH RELEVANCE: As the underlying cause of heart attack and stroke, atherosclerosis (plaque formation in arteries) is the most common cause of death and suffering in the industrialized world. Macrophage apoptosis (programmed cell death) has been implicated in the development of atherosclerotic lesions and plaque rupture, which results in clot formation and arterial occlusion and heart attack. The goal of these studies is to define the roles of two key signaling proteins (Akt and IKKalpha) in macrophage survival and atherosclerosis, which will help define new targets for the prevention and treatment of atherosclerosis.